Hibernating ground squirrels have dramatically decreased heart rates and blood flow which should put them at risk of forming blood clots. In response, they have several adaptations that decrease blood clotting, including a decrease in the levels of blood clotting factors VIII and IX, and a 90% decrease in platelets. The platelet decrease is due to reorganization of microtubules leading to a conformational change in the platelets causing them to be trapped in the spleen. Within two hours of arousal in the spring, platelet levels and blood clotting times return to normal. The specific aims of this proposal are to study the effects of hibernation on platelet activity, the regulation of blood clotting factors, and fibrinolysis, or the breakdown of fibrin clots. Platelet activity will be measured before, during an immediately after hibernation by detection of markers of activation on the surface of platelets and by platelet aggregation and adhesion. Human platelets that are stored in the cold are rapidly cleared by the liver when re-injected back into a patient. As ground squirrel platelets are essentially stored in the cold for six months while hibernating, we will test if this same mechanism occurs by injecting ground squirrels with warmed and chilled platelets and following their clearance in vivo. This could have direct application in extending storage of human platelets in the cold. Hibernation also reversibly reduces the activity of blood clotting factors VIII and IX, but not most of the other clotting factors. Understanding how these two clotting factors activities are reversibly regulated could lead to improved treatments of hemophilia A and B. Several other proteins that are known to regulate blood clotting have not been studied in hibernating animals, and could be additional candidate proteins to regulate blood clotting during periods of low blood flow. Finally, the impact of hibernation on the breakdown of blood clots has not been measured, and we will compare ground squirrel plasma before, during and immediately after hibernation for markers of fibrinolysis, like D-dimer, and the activity of proteins involved in fibrinolysis. This research will improve our understanding of how a mammal can adapt to reduce the risk of blood clots during low blood flow, and preserve platelet activity after hypothermia. These findings could have direct clinical applications in allowing human platelets to be stored in the cold, regulating blood coagulation in cases of accidental or induced hypothermia, and reversible regulation of factor VIII and IX activity. PUBLIC HEALTH RELEVANCE: This research could identify ways to store human platelets in the cold which would increase their shelf life. The research could also lead to treatments that reversibly regulate two important blood clotting factors, possibly as long-term anti-clotting therapy. [unreadable] [unreadable] [unreadable] [unreadable]